Data from recent studies has provided highly suggestive evidence that glucagon is synthesized via a biosynthetic precursor (proglucagon). However, proglucagon has not been isolated and chemically characterized from any species. Glucagon immunoreactive peptides larger than glucagon in molecular size which incorporate 3H-tryptophan during in vitro incorporation studies have been extracted from pancreas or pancreatic islets of teleost fish, birds and several mammalian species including humans. From our studies using anglerfish islets, we have identified on the basis of several criteria a peptide having a molecular size of approximately 12,000 daltons as anglerfish "proglucagon". Cell-free translation of anglerfish islet mRNA generates four major translation products, all of which are less than 18,000 daltons in molecular size. Yet when whole tissue is extracted directly in acetic acid, glucagon immunoreactive peptides greater than 20,000 daltons are found. One goal of the proposed research is to define further the role (if any) of the peptides with m.w. greater than 20,000 in glucagon biosynthesis by employing continuous and pulse-chase incubations to label them. Using gel filtration, ion exchange chromatography, and bi-dimensional isofocusing-polyacrylamide gel electrophoresis, attempts will be made to isolate and purify the 12,000 m.w. and larger peptides. When sufficient quantities are recovered, chemical analyses including N- and C- terminal analysis, amino acid composition determination and partial sequence determination will be made. An additional goal is to identify by immunologic means which of the cell-free translation products is glucagon-related and to define further its role in the biosynthesis of glucagon.